382. LARGE SCALE PRODUCTION OF LANGERHANS CELLS/PLASMACYTOID DENDRITIC CELLS FOR ALLERGENICITY STUDIES AND IN VITRO TISSUE ENGINEERING.

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Ayehunie, S., Lamore, S., Snell, M., Klausner, M., and Sheasgreen, J. MatTek Corporation, Ashland, MA. (USA). Presented at 5th World Congress, Berlin, Germany, August, (2005). Presented at 9th International Workshop on Langerhans Cells, Madeira, Portugal (2005).

Summary:
This study by MatTek scientists demonstrated that MatTek’s Human Dendritic Cells are an excellent in vitro model for use in a wide variety of studies related to allergenicity, microbial infection and transmission, neutralizing antibodies and anti-microbial agents, antigen capture and presentation, innate immunity and immuno-therapy.

Langerhans cells (LC)/dendritic cells (DC) residing in mucosal tissues play a key role in sensitization and viral and bacterial infections.

Recently, MatTek scientists have developed an efficient way of generating large numbers of DC from human umbilical cord blood-derived CD34+ progenitor cells. As a result, some of the limiting factors for a wider use of DC in biological research have been overcome.

The improved method resulted in the generation of an average of >400 million DC from 1 million starting progenitor CD34+ cells.

The DC express CD1a, HLA-DR, CD80, CD86, CD40, CD54, CD11c, and CD206 (mannose receptor), and unlike monocyte derived DC, the DC express Birbeck granules.

The lifespan of the DC has been increased to 29 days with detectable Birbeck granules and very small phenotypic changes over this period.

When compared with unstimulated controls, lipopolysaccharide (LPS) plus TNF-a stimulated the DC to adopt a more mature phenotype. Increased mean fluorescence intensity (MFI) for HLA-DR (7 fold), CD83 (3 fold), and CD86 (4 fold) and released IL-12 (94 fold) and RANTES (202 fold) were observed.

The non-stimulated DC can be fractionated by FACS sorting into plasmacytoid DC (pDC; CD123+CD11c-) and myeloid DC (mDC, CD123-CD11c+). Compared to the mDC, the pDC have shorter dendrites, express lower level of CD1a (11.8% vs. 66.8%), and can be expanded 6-15 fold using specially formulated medium (mDC do not proliferate). The pDC also showed increased levels of CD86 surface expression following treatment with the allergens (n=7), but not following exposure to irritants (n=5).

The DC can also be incorporated into in vitro reconstructed three-dimensional, epidermal, oral, and ectocervical-vaginal tissue models for sensitization and HIV-1 infection studies.

In conclusion, the generated DC alone or the DC containing in vitro engineered tissue models will likely be useful in a variety of studies related to:
1) allergenicity,
2) microbial infection and transmission,
3) neutralizing antibodies and anti-microbial agents,
4) antigen capture and presentation,
5) innate immunity, and
6) immuno-therapy.

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Dendritic Cells Data Sheet

Dendritic Cells Specifications

Dendritic Cells Technical References

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Applications: Allergens/Allergenicity, Antigen presentation, Characterization study, Immunological research, Innate immunity

Keywords: Allergenicity, Antigen-presenting cells (APC), Birbeck granules, CD11c, CD1a, CD206, CD209, CD34, CD40, CD54, CD80, CD86, Chemokine, Cytokine, Dendritic cells (DC), FACS sorting, Green fluorescent protein (GFP), HIV-1, HLA-DR, IL-12, In Vitro, Langerhans cells (LC), Mean fluorescence intensity (MFI), Myeloid dendritic cells, PDC, Plasmacytoid, Plasmacytoid dendritic cells, RANTES, TNF-a, Tissue engineering, Transmission electron microscopy, pDC

Endpoints: CD 206, CD1a, CD40, CD54, CD86, HLA-DR, IL-12, RANTES

Materials Tested: Balsam of peru, Bandowski’s base, Cinnamaldehyde, Ethanol, Fluorescein-5-isothiocyanate (FITC), Hydroquinone, Lipopolysaccharide (LPS), Neomycin sulfate, Nickel sulfate, Para-phenylene diamine, Potassium hydroxide, Propylene glycol, Sodium dodecyl sulfate (SDS), Tween 20

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